The purpose of this project is to develop vaccines against otitis media caused by nontypeable Haemophilus influenzae and Moraxella catarrhalis. In this fiscal year, efforts are focusing on investigating if intranasal immunization with a detoxified lipooligosaccharide (LOS)-CRM (cross-reactive mutant of diphtheria toxin) vaccine candidate would generate protective immunity against M. catarrhalis in a mouse model of pulmonary clearance. Intranasal immunization with dLOS-CRM vaccine candidate plus cholera toxin induced a significantly dose-dependent increase of IgA and IgG in the nasal wash, lung lavage, saliva, and fecal extract. In addition, serum IgG, IgM, and IgA against LOS of M. catarrhalis were detected. LOS-specific antibody-forming cells were found in the nasal passages, spleens, nasal-associated lymphoid tissues, cervical lymph nodes, lungs, and Peyer's patches using an enzyme-linked immunospot assay. The dLOS-CRM vaccine induced a significant bacterial clearance (70-90%) of both homologous and heterologous strains in the lungs compared to the controls (p value less than 0.01). Intriguingly, intranasal immunization with dLOS-CRM showed a higher level of bacterial clearance compared with subcutaneous injections with dLOS-CRM. These data indicate that dLOS-CRM induces specific mucosal and systemic immunity through intranasal immunization, and also provides effective bacterial clearance. On the basis of these results, we believe that dLOS-CRM vaccine candidate should undergo continued testing to determine whether it would induce protective immune response in humans.